Many recent research efforts have examined the role that saturated and unsaturated fatty acids play in reducing the risk of coronary heart disease. In the past, it was believed that monounsaturates, in contrast to saturates and polyunsaturates, had no effect on serum cholesterol and coronary heart disease risk. Several recent human clinical studies suggest that diets high in monounsaturated fat and low in saturated fat may reduce the "bad" (low-density lipoprotein) cholesterol while maintaining the "good" (high-density lipoprotein) cholesterol (Mattson et al. Journal of Lipid Research, (1985) 26:194-202; herein incorporated by reference).
Vegetable oils may play an important role in shifting the balance towards production of "good" cholesterol. The specific performance and health attributes of edible oils is determined largely by their fatty acid composition. Most vegetable oils derived from commercial varieties are composed primarily of palmitic (16:0), stearic (18:0), oleic (18:1), linoleic (18:2) and linolenic (18:3) acids. Palmitic and stearic acids are, respectively, 16- and 18-carbon-long, saturated fatty acids. Oleic, linoleic and linolenic are 18-carbon-long, unsaturated fatty acids containing one, two and three double bonds, respectively. Oleic acid is referred to as a monounsaturated fatty acid, while linoleic and linolenic acids are referred to as polyunsaturated fatty acids.
The relative amounts of saturated and unsaturated fats in commonly used edible vegetable oils are summarized below (Table 1):
TABLE 1 ______________________________________ Percentages of Saturated and Unsaturated Fatty Acids in the Oils of Selected Oil Crops Saturated Monounsaturated Polyunsaturated ______________________________________ Canola 6% 58% 36% Soybean 15% 24% 61% Corn 13% 25% 62% Peanut 18% 48% 34% Safflower 9% 13% 78% Cotton 30% 19% 51% ______________________________________
A vegetable oil low in total saturates and high in monounsaturate would provide significant health benefits to consumers as well as economic benefits to oil processors. Soybean and corn varieties which produce seeds containing such an improved oil would also produce valuable meal as animal feed.
Another type of desirable vegetable oil is a substitute for palm oil and its fractionation products. Palm oil is the world's second most important vegatable oil, after soybean oil (Gascon et al., Oil Palm, In Oil Crops of the World, Robbelen et al., Ed., (1989) McGraw-Hill, Chapter 27). About 80% of the world palm oil is supplied by Malaysia and Indonesia, the remainder coming from Africa and South America. Palm oil is widely used in the manufacture of hardened vegetable fats such as margarines and shortenings. Palm stearin, about 10% of the palm oil, is used as a hardstock to increase the creaming properties of margarine blends and whipped toppings (Traitlet et al., J. Amer. Oil Chemists Soc., (1985) 62:417-421). Both palm oil and palm stearin have well-known non-food uses in the manufacture of soaps and lubricating oils. Palm olein (60% of the oil) is useful for cocking oils and interesterification of palm stearin with palm olein provides a hardstock for spread formulations without the use of hydrogenated fat components. Finally, the Palm Mid-Fraction (PMF), which is palm oil minus the palm stearin and palm olein fractions, is suitable for the manufacture of cocoa-butter substitutes (Traitler et al., J. Amer. Oil Chemists Soc., (1985), 62:417-421). Commercial palm oil con,sins 44% palmitate (P), 4.5% stearate (S) and 39.2% oleate (O) (Gunstone et al., The Lipid Handbook, Chapman-Hall, (1986) 176). Palm stearin (47-74% P, 4.4-5.6% S, 15.6-37% O), palm olein (39.8% P, 4.4% S, 42.5% O) and PMF (43 % P, 5.6% S, 24.3% O) are produced by fractionation of palm oil (Gunstone et al., The Lipid Handbook, Chapman-Hall, (1986) 178). Thus, a vegetable oil, such as soybean, with an increased level palmitic acid, especially in oilseed lines containing suitable levels of oleic acid and reduced levels of unsaturated fatty acids, could yield a substitute for palm oil. Such a soybean or other oil seed could also yield a substitute for palm stearin, clein and PMF without the need for costly fractionation procedures. This would add value to oil and food processors as well as reduce the foreign import of palm oil.
Oil biosynthesis in plants has been fairly well-studied (Browse et al., Ann. Rev. Plant Physiol. Mol. Biol. (1991) 42: 467-506; Science (1991) 252:80-87)), From these studies it is apparent that in seed tissue the rate-limiting step in the metabolism of palmitic acid to stearic acid and the subsequent formation of oleic acid is the reaction catalyzed by the enzyme .beta.-ketoacyl-ACP synthetase II. Thus, .beta.-ketoacyl-ACP synthetase II is an attractive target for modification by genetic engineering since a decrease in .beta.-ketoacyl-ACP synthetase II activity would presumably lead to an increased palmitic acid content of the plant oil and an increase in .beta.-ketoacyl-ACP synthetase II activity would presumably lead to higher unsaturated fatty acids at the expense of saturated fats in the oil.
No evidence exists in the public art that complete isolation of a plant .beta.-ketoacyl-ACP synthetase II has been accomplished. The partial purification of a .beta.-ketoacyl-ACP synthetase II was reported from spinach leaves (Shimakata et al., Proc. Natl. Acad. Sci. (1982) 79:5808-5812) and oilseed rape (MacKintosh et al., Biochim. Biophys. Acta. (1989) 1002:114-124) but in neither case was the purification sufficient to identify a single protein associated with .beta.-ketoacyl-ACP synthetase II activity. Furthermore, there is no evidence that a method to control the levels of saturated and unsaturated fatty acids in edible plants is known in the art.